Shabih U. Hasan

Respiratory Control in Neonatal Rats Exposed to Prenatal Cigarette Smoke

RATIONALE Prenatal cigarette smoke (CS) exposure, increased environmental temperature, and hypoxic episodes have been postulated as major risk factors for sudden infant death syndrome. OBJECTIVES To test the hypothesis that maternal CS exposure disrupts eupneic breathing and depresses breathing responses of neonatal rats to thermal and hypoxic challenges. METHODS Experiments were performed on 1-week-old rat pups exposed prenatally to CS (n = 39) or room air (sham; n = 30). Breathing patterns were recorded by whole-body plethysmography during thermoneutral or hyperthermic states under normoxic and hypoxic conditions. MEASUREMENTS AND MAIN RESULTS Mean pup weight, breaths per minute, and gasping respiratory patterns were measured for both smoke- and sham-exposed groups during thermoneutral and hyperthermic states under normoxic and hypoxic conditions. Under thermoneutral conditions, hypoxia caused gasping in CS-exposed animals but not in sham-exposed animals. Furthermore, under hyperthermic conditions, whereas hypoxia induced gasping in both groups, only CS-exposed animals exhibited a pronounced and longer lasting respiratory depression after the termination of hypoxia. CONCLUSIONS We show that prenatal CS exposure increases the likelihood of gasplike respiration and provide the first experimental evidence that the combined effects of prenatal CS exposure and hyperthermia dramatically prolong the time required for neonates to return to eupneic breathing after hypoxia. These observations provide important evidence of how prenatal CS exposure, hypoxic episodes, and hyperthermia might place infants at higher risk for sudden infant death syndrome.

Prenatal cigarette smoke exposure selectively alters protein kinase C and nitric oxide synthase expression within the neonatal rat brainstem.

Maternal smoking is a major risk factor for sudden infant death syndrome. Protein kinase C (PKC) and neuronal nitric oxide synthase (NOS) activities within the dorsocaudal brainstem (DB) mediate critical components of respiratory drive and could be implicated in SIDS. Thus, exposure to smoking during fetal life could modify the expression of these kinases in the DB. Rats were exposed to cigarette smoke or room air (Sham) from day 2 to 22 of pregnancy. Immunoblots of DB lysates at 2 days postnatally revealed no differences in PKC-alpha, PKC-beta, and endothelial NOS expression. However, PKC-gamma, PKC-delta, and neuronal NOS immunoreactivities were reduced in the cigarette smoke group. We conclude that gestational smoking is associated with selective reductions in PKC and NOS isoforms within the DB, which could decrease respiratory drive and lead to enhanced hypoxic vulnerability in infants of smoking mothers.

Target cell contact suppresses neurite outgrowth from soma–soma paired Lymnaea neurons

Neurite extension from developing and/or regenerating neurons is terminated on contact with their specific synaptic partner cells. However, a direct relationship between the effects of target cell contact on neurite outgrowth suppression and synapse formation has not yet been demonstrated. To determine whether physical/synaptic contacts affect neurite extension from cultured cells, we utilized soma-soma synapses between the identified Lymnaea neurons. A presynaptic cell (right pedal dorsal 1, RPeD1) was paired either with its postsynaptic partner cells (visceral dorsal 4, VD4, and Visceral dorsal 2, VD2) or with a non-target cell (visceral dorsal 1, VD1), and the interactions between their neurite outgrowth patterns and synapse formation were examined. Specifically, when cultured in brain conditioned medium (CM, contains growth-promoting factors), RPeD1, VD4, and VD2 exhibited robust neurite outgrowth within 12-24 h of their isolation. Synapses, similar to those seen in vivo, developed between the neurites of these cells. RPeD1 did not, however, synapse with its non-target cell VD1, despite extensive neuritic overlap between the cells. When placed in a soma-soma configuration (somata juxtaposed against each other), appropriate synapses developed between the somata of RPeD1 and VD4 (inhibitory) and between RPeD1 and VD2 (excitatory). Interestingly, pairing RPeD1 with either of its synaptic partner (VD4 or VD2) resulted in a complete suppression of neurite outgrowth from both pre- and postsynaptic neurons, even though the cells were cultured in CM. A single cell in the same dish, however, extended elaborate neurites. Similarly, a postsynaptic cell (VD4) contact suppressed the rate of neurite extension from a previously sprouted RPeD1. This suppression of the presynaptic growth cone motility was also target cell contact specific. The neurite suppression from soma-soma paired cells was transient, and neuronal sprouting began after a delay of 48-72 h. In contrast, when paired with VD1, both RPeD1 and this non-target cell exhibited robust neurite outgrowth. We demonstrate that this neurite suppression from soma-soma paired cells was target cell contact/synapse specific and Ca(2+) dependent. Specifically, soma-soma pairing in CM containing either lower external Ca(2+) concentration (50% of its control level) or Cd(2+) resulted in robust neurite outgrowth from both cells; however, the incidence of synapse formation between the paired cells was significantly reduced. Taken together, our data show that contact (physical and/or synaptic) between synaptic partners strongly influence neurite outgrowth patterns of both pre- and postsynaptic neurons in a time-dependent and cell-specific manner. Moreover, our data also suggest that neurite outgrowth and synapse formation are differentially regulated by external Ca(2+) concentration.

An identified central pattern-generating neuron co-ordinates sensory-motor components of respiratory behavior in Lymnaea.

Defining the attributes of individual central pattern‐generating (CPG) neurons underlying various rhythmic behaviors are fundamental to our understanding of how the brain controls motor programs, such as respiration and locomotion. To this end, we have explored a simple invertebrate preparation in which the neuronal basis of respiratory rhythmogenesis can be investigated from the whole animal to a single cell level. An identified dopaminergic neuron, termed right pedal dorsal 1 (RPeD1), is a component of the CPG network which controls hypoxia‐driven, aerial respiration in the fresh water snail Lymnaea stagnalis. Using intact, semi‐intact and isolated brain preparations, we have discovered that in addition to its role as a respiratory CPG neuron, RPeD1 co‐ordinates sensory‐motor input from the pneumostome (the respiratory orifice) at the water/air interface to initiate respiratory rhythm generation. An additional, novel role of RPeD1 was also found. Specifically, direct intracellular stimulation of RPeD1 induced pneumostome openings, in the absence of motor neuronal activity. To determine further the role of RPeD1 in the respiratory behavior of intact animals, either its axon was severed or the soma selectively killed. Many components of the respiratory behavior in the intact animals were found to be perturbed following RPeD1 axotomy or ‘somatomy’ (soma removed). Taken together, the data presented provide a direct demonstration that RPeD1 is a multifunctional CPG neuron, which also serves many additional roles in the control of breathing behavior, ranging from co‐ordination of mechanosensory input to the motor control of the respiratory orifice.

Significance of vagal innervation in perinatal breathing and gas exchange.

The mechanisms responsible for the establishment of continuous breathing at birth remain unknown. Several studies have shown that postnatal vagal denervation produces deleterious effects on ventilation as well as breathing patterns during the newborn period. However, the validity of these studies was compromised by anesthesia, tracheostomy or possible secondary laryngeal obstruction. We have recently developed an unanesthetized lamb model in which vagal denervation was performed antenatally and below the recurrent laryngeal nerves thereby avoiding anesthesia, tracheostomy and laryngeal paralysis. The denervated animals developed life-threatening respiratory failure shortly after birth, implying that vagal innervation of the lungs plays an essential role in establishing adequate gas exchange in the first hours after birth. We have subsequently investigated various mechanisms of respiratory failure in denervated animals. Our results show that the surfactant system dysfunction and loss of vagally mediated volume feedback likely contributed to the respiratory failure observed in the intrathoracically denervated animal model.

Prostaglandins are responsible for the inhibition of breathing observed with a placental extract in fetal sheep

We have previously observed that the infusion of a placental extract inhibits breathing movements in fetal sheep, suggesting that a placental factor may be responsible for the inhibition of fetal breathing. Our preliminary results suggested that a small peptide or a substance bound to a peptide was likely responsible for this inhibition. Since prostaglandins are found in high concentrations in the placenta, it is possible that they may be responsible for the inhibition of breathing observed with the placental extract. We hypothesized that if prostaglandins were the active factors in the placental extract, then inhibition of the production of placental prostaglandins should eliminate the activity of the extract. We infused untreated and indomethacin/ASA-treated placental extracts into the carotid artery of eight chronically instrumented fetal sheep continuously over 3 h. The concentration of all prostaglandins measured in the untreated placental extracts were significantly higher than in the indomethacin/ASA-treated extracts. Only the infusion of the untreated placental extract induced a significant decreased in the incidence of fetal breathing. Fetal plasma prostaglandins increased significantly only with the infusion of the untreated placental extracts. These findings suggest that the inhibition of breathing observed with the placental extract is likely related to prostaglandins.

Neonatal Ventricular Tachyarrhythmias in Medium Chain Acyl-CoA Dehydrogenase Deficiency

We describe an unusually severe case of medium chain acyl-CoA dehydrogenase (MCAD) deficiency in a term female neonate, who presented at 12 h of age with lethargy, poor feeding, hypoglycemia and ventricular tachyarrhythmias. While arrhythmias are common in other disorders of fatty acid beta-oxidation, ventricular tachyarrhythmias have rarely been reported with MCAD deficiency in childhood. Since the results of newborn metabolic screening are usually not available within the first 3 days of life, our case highlights the need for health care professionals to be made aware of this early and uncommon but potentially fatal presentation of MCAD deficiency.

Intratracheal Administration of Budesonide‐Surfactant in Prevention of Bronchopulmonary Dysplasia in Very Low Birth Weight Infants: A Systematic Review and Meta‐Analysis

Despite the near universal adaptation of gentle mechanical ventilation, surfactant use and non‐invasive respiratory support, bronchopulmonary dysplasia (BPD) remains one of the most common respiratory morbidities in very low birth weight (VLBW) infants. Thus, the objective of this review was to evaluate the efficacy of intra‐tracheal administration of budesonide‐surfactant mixture in preventing bronchopulmonary dysplasia (BPD) in very low birth weight (VLBW) infants. MEDLINE, EMBASE, and PubMed were searched for randomized clinical trials in which intra‐tracheal administration of budesonide‐surfactant was used to prevent BPD in infants. The primary outcomes were BPD and composite outcome of death or BPD. Meta‐analysis of the two clinical trials revealed that infants who received intra‐tracheal instillation of budesonide‐surfactant mixture demonstrated 43% reduction in the risk of BPD (RR: 0.57; 95%CI: 0.43‐0.76, NNT = 5). Although mortality was not different between the groups (OR: 0.61; 95%CI: 0.34‐1.04), a 40% reduction was observed in the composite outcome of death or BPD in the budesonide‐surfactant group (RR: 0.60; 95%CI: 0.49‐0.74, NNT = 3). Thus, this review concludes that intra‐tracheal administration of budesonide‐surfactant combination was associated with decreased incidence of BPD alone or composite outcome of death or BPD in VLBW infants though there is a need for larger trials before it can be recommended as a standard of care.

Bilateral cataracts associated with glucose-6-phosphate dehydrogenase deficiency

Glucose-6-phosphate dehydrogenase (G6PD) has an essential role in the defense against cellular oxidative injury. In neonates, the most common manifestation of G6PD deficiency is jaundice and hemolysis due to factors causing oxidative stress. Less known are the ocular associations described with G6PD deficiency, including cataracts. Oxidative injury is involved in the pathogenesis of almost all forms of cataracts, causing the lens proteins to undergo modifications, denaturation and form insoluble aggregates resulting in cataracts. Although cataracts in adult males have been reported in several studies, there are few reports of cataracts in infants with G6PD deficiency. We describe a preterm male neonate with G6PD deficiency who developed bilateral cataracts following an episode of neonatal sepsis and severe hemolysis necessitating an exchange blood transfusion.

Role of vagal innervation on pulmonary surfactant system during fetal development

Vagally mediated afferent feedback and compliant lungs (surfactant system) play vital roles in the establishment of adequate alveolar ventilation and pulmonary gas exchange at birth. Although the significance of vagal innervation in the establishment of normal breathing patterns is well recognized, the precise role of lung innervation in the maturation of the surfactant system remains unclear. The specific aim of the present study was to investigate whether vagal denervation compromises the surfactant system during fetal development. Experiments were performed on 12 time-dated fetal sheep: 8 underwent cervical vagal denervation, and 4 were sham operated. Vagal denervation was performed at 110-113 days gestation. Fetal lambs were instrumented in utero to record arterial pH and blood-gas tensions. The animals were delivered by cesarean section under general anesthesia between 130 and 133 days gestation (term approximately 147 days). Lung samples were collected for wet-to-dry ratios, light and electron microscopy, and overall lung morphology. In addition, total proteins, total phospholipids, and surfactant proteins A and B were analyzed in both lung tissue and bronchoalveolar lavage fluid. Vagal denervation had no effect on alveolar architecture, including type II cells or the morphology of lamellar bodies within them. Furthermore, surfactant proteins A and B and total phospholipids were similar in lung tissue and bronchoalveolar lavage fluid between the two groups. A significant correlation was observed between circulating cortisol concentrations and surfactant proteins in the bronchoalveolar lavage fluid and lung tissue. We provide definitive evidence that vagal innervation at midgestation is not required for maturation of the pulmonary surfactant system during fetal development.